No abstract

Fructo-oligosaccharides (FOS) are linear fructans comprising 2–5 fructose units linked to a terminal glucose residue. They are widely used as food and feed additives due to their sweetness, low calorific value, and prebiotic properties. Here we describe the synthesis of FOS catalyzed by a cell-free crude enzyme solution containing recombinant fructosyltransferase (1-FFT) produced in the yeast Kluyveromyces lactis. During the enzyme catalysis, glucose accumulates as a by-product and eventually inhibits FOS production. We therefore used an enzyme membrane reactor (EMR) to achieve the continuous removal of glucose and the simultaneous replenishment of sucrose. We observed a loss of flux during the reaction with the characteristics of complete pore blocking, probably caused by a combination of proteins (enzyme molecules) and polysaccharides (FOS). Such complex fouling mechanisms must be overcome to achieve the efficient production of FOS using EMR systems.

Fructo-oligosaccharides (FOS) are prebiotic low-calorie sweeteners that are synthesized by the transfer of fructose units from sucrose by enzymes known as fructosyltransferases. If these enzymes generate β-(2,6) glycosidic bonds, the resulting oligosaccharides belong to the neoseries (neoFOS). Here, we characterized the properties of three different fructosyltransferases using a design of experiments approach based on response surface methodology with a D-optimal design. The reaction time, pH, temperature, and substrate concentration were used as parameters to predict three responses: The total enzyme activity, the concentration of neoFOS and the neoFOS yield relative to the initial concentration of sucrose. We also conducted immobilization studies to establish a cascade reaction for neoFOS production with two different fructosyltransferases, achieving a total FOS yield of 47.02 ± 3.02%. The resulting FOS mixture included 53.07 ± 1.66 mM neonystose (neo-GF3) and 20.8 ± 1.91 mM neo-GF4.